1. Field of the Invention
The invention relates to linear cyclodextrin copolymers and linear oxidized cyclodextrin copolymers. These copolymers, respectively, contain a cyclodextrin moiety, unoxidized or oxidized, as a monomer unit integrated into the copolymer backbone. The invention also relates methods of preparing linear cyclodextrin copolymers and linear oxidized cyclodextrin copolymers. Such cyclodextrin copolymers may be used as a delivery vehicle of various therapeutic agents.
2. Background of the Invention
Cyclodextrins are cyclic polysacchaides containing naturally occurring D(+)-glucopyranose units in an xcex1-(1,4) linkage. The most common cyclodextrins are alpha (xcex1)-cyclodextrins, beta (xcex2)-cyclodextrins and gamma (xcex3)-cyclodextrins which contain, respectively. six, seven or eight glucopyranose units. Structurally, the cyclic nature of a cyclodextrin forms a torus or donut-like shape having an inner apolar or hydrophobic cavity, the secondary hydroxyl groups situated on one side of the cyclodextrin torus and the primary hydroxyl groups situated on the other. Thus, using (xcex2)-cyclodextrin as an example, a cyclodextrin is often represented schematically as follows: 
The side on which the secondary hydroxyl groups are located has a wider diameter than the side on which the primary hydroxyl groups are located. The hydrophobic nature of the cyclodextrin inner cavity allows for the inclusion of a variety of compounds. (Comprehensive Supramolecular Chemistry, Volume 3, J. L. Atwood et al., eds., Pergamon Press (1996); T. Cserhati, Analytical Biochemistry, 225:328-332 (1995); Husain et al., Applied Spectroscopy, 46:652-658 (1992); FR 2 665 169).
Cyclodextrins have been used as a delivery vehicle of various therapeutic compounds by forming inclusion complexes with various drugs that can fit into the hydrophobic cavity of the cyclodextrin or by forming non-covalent association complexes with other biologically active molecules such as oligonucleotides and derivatives thereof. For example, U.S. Pat. No. 4,727,064 describes pharmaceutical preparations consisting of a drug with substantially low water solubility and an amorphous, water-soluble cyclodextrin-based mixture. The drug forms an inclusion complex with the cyclodextrins of the mixture. In U.S. Pat. No. 5,691,316, a cyclodextrin cellular delivery system for oligonucleotides is described. In such a system, an oligonucleotide is noncovalently complexed with a cyclodextrin or, alternatively, the oligonucleotide may be covalently bound to adamantine which in turn is non-covalently associated with a cyclodextrin.
Various cyclodextrin containing polymers and methods of their preparation are also known in the art. (Comprehensive Supramolecular Chemistry, Volume 3, J. L. Atwood et al., eds., Pergamon Press (1996)). A process for producing a polymer containing immobilized cyclodextrin is described in U.S. Pat. No. 5,608,015. According to the process, a cyclodextrin derivative is reacted with either an acid halide monomer of an xcex1,xcex2-unsaturated acid or derivative thereof or with an xcex1,xcex2-unsaturated acid or derivative thereof having a terminal isocyanate group or a derivative thereof. The cyclodextrin derivative is obtained by reacting cyclodextrin with such compounds as carbonyl halides and acid anhydrides. The resulting polymer contains cyclodextrin units as side chains off a linear polymer main chain.
U.S. Pat. No. 5,276,088 describes a method of synthesizing cyclodextrin polymers by either reacting polyvinyl alcohol or cellulose or derivatives thereof with cyclodextrin derivatives or by copolymerization of a cyclodextrin derivative with vinyl acetate or methyl methacrylate. Again, the resulting cyclodextrin polymer contains a cyclodextrin moiety as a pendant moiety off the main chain of the polymer.
A biodegradable medicinal polymer assembly with supermolecular structure is described in WO 96/09073 A1. The assembly comprises a number of drug-carrying cyclic compounds prepared by binding a drug to an xcex1, xcex2, or xcex3-cyclodextrin and then stringing the drug/cyclodextrin compounds along a linear polymer with the biodegradable moieties bound to both ends of the polymer. Such an assembly is reportably capable of releasing a drug in response to a specific biodegradation occurring in a disease. These assemblies are commonly referred to as xe2x80x9cnecklace-typexe2x80x9d cyclodextrin polymers.
However, there still exists a need in the art for linear cyclodextrin polymers in which the cyclodextrin moiety is part of the main chain and not a pendant moiety off the main chain and a method for their preparation.
This invention answers this need by providing a linear cyclodextrin copolymer. Such a linear cyclodextrin copolymer has a repeating unit of formula Ia, Ib, or a combination thereof: 
The invention also provides methods of preparing a linear cyclodextrin copolymer. One method copolymerizes a cyclodextrin monomer precursor disubstituted with the same or different leaving group and a comonomer A precursor capable of displacing the leaving group. Another such method involves iodinating a cyclodextrin monomer precursor to form a diiodinated cyclodextrin monomer precursor and then copolymerizing the diiodinated cyclodextrin monomer precursor with a comonomer A precursor to produce the linear cyclodextrin copolymer. Another method involves iodinating a cyclodextrin monomer precursor to form a diiodinated cyclodextrin monomer precursor, aminating the diiodinated cyclodextrin monomer precursor to form a diaminated cyclodextrin monomer precursor and then copolymerizing the diaminated cyclodextrin monomer precursor with a comonomer A precursor to produce the linear cyclodextrin copolymer. Yet another method involves the reduction of a linear oxidized cyclodextrin copolymer to the linear cyclodextrin copolymer.
The invention further provides a linear oxidized cyclodextrin copolymer. A linear oxidized cyclodextrin copolymer is a linear cyclodextrin copolymer which contains at least one oxidized cyclodextrin moiety of formula VIa or VIb: 
Each cyclodextrin copolymer of the invention may be oxidized so as to form a linear oxidized cyclodextrin copolymer having a repeating unit of formula VIa, VIb, or a combination thereof.
The invention also provides a method of preparing a linear oxidized cyclodextrin copolymer. One method involves oxidizing a linear cyclodextrin copolymer such that at least one cyclodextrin monomer is oxidized. Other methods involve copolymerizing an oxidized cyclodextrin monomer precurser with a comonomer A precursor.
The invention still further provides a linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer grafted onto a substrate and a method of their preparation. The invention also provides a linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer crosslinked to another polymer and a method of their preparation. A method of preparing crosslinked cyclodextrin polymers involves reacting a linear or linear oxidized cyclodextrin copolymer with a polymer in the presence of a crosslinking agent.
The invention provides a linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer having at least one ligand bound to the cyclodextrin copolymer. The ligand may be bound to either the cyclodextrin moiety or the comonomer A moiety of the copolymer.
The invention also provides a cyclodextrin composition containing at least one linear cyclodextrin copolymer of the invention and at least one linear oxidized cyclodextrin copolymer of the invention. The invention also provides therapeutic compositions containing a therapeutic agent and a linear cyclodextrin copolymer and/or a linear oxidized cyclodextrin copolymer of the invention. A method of treatment by administering a therapeutically effective amount of a therapeutic composition of the invention is also described.
FIG. 1 depicts Transfection Studies with Plasmids Encoding Luciferase Reporter Gene: FIG. 1A, Transfection with copolymer 16; and FIG. 1B, Toxicity of copolymer 16 to BHK-21.
One embodiment of the invention is a linear cyclodextrin copolymer. A linear cyclodextrin copolymer is a polymer containing cyclodextrin moieties as an integral part of its polymer backbone. Previously, cyclodextrin moieties were not a part of the main polymer chain but rather attached off a polymer backbone as pendant moieties.
According to the invention, a linear cyclodextrin copolymer has a repeating unit of formula Ia, Ib, or a combination thereof: 
In formula Ia and Ib, C is a substituted or unsubstituted cyclodextrin monomer and A is a comonomer bound, i.e. covalently bound, to cyclodextrin C. Polymerization of a cyclodextrin monomer C precursor with a comonomer A precursor results in a linear cyclodextrin copolymer of the invention. Within a single linear cyclodextrin copolymer of the invention, the cyclodextin monomer C unit may be the same or different and, likewise, the comonomer A may be the same or different.
A cyclodextrin monomer precursor may be any cyclodextrin or derivative thereof known in the art. As discussed above, a cyclodextrin is defined as a cyclic polysaccharide most commonly containing six to eight naturally occurring D(+)-glucopyranose units in an xcex1-(1,4) linkage. Preferably, the cyclodextrin monomer precursor is a cyclodextrin having six, seven and eight glucose units, i.e., respectively, an alpha (xcex1)-cyclodextrin, a beta (xcex2)-cyclodextrin and a gamma (xcex3)-cyclodextrin. A cyclodextrin derivative may be any substituted cyclodextrin known in the art where the substituent does not interfere with copolymerization with comonomer A precursor as described below. According to the invention, a cyclodextrin derivative may be neutral, cationic or anionic. Examples of suitable substituents include, but are not limited to, hydroxyalkyl groups, such as, for example, hydroxypropyl, hydroxyethyl; ether groups, such as, for example, dihydroxypropyl ethers, methyl-hydroxyethyl ethers, ethyl-hydroxyethyl ethers, and ethyl-hydroxypropyl ethers; alkyl groups, such as, for example, methyl; saccharides, such as, for example, glucosyl and maltosyl; acid groups, such as, for example, carboxylic acids, phosphorous acids, phosphinous acids, phosphonic acids, phosphoric acids, thiophosphonic acids, thiophosphonic acid and sulfonic acids; imidazole groups; and sulfate groups.
A cyclodextrin monomer precursor may be further chemically modified (e.g. halogenated, aminated) to facilitate or affect copolymerization of the cyclodextrin monomer precursor with a comonomer A precursor, as described below. Chemical modification of a cyclodextrin monomer precursor allows for polymerization at only two positions on each cyclodextrin moiety, i.e. the creation of a bifunctional cyclodextrin moiety. The numbering scheme for the C1-C6 positions of each glucopyranose ring is as follows: 
In a preferred embodiment, polymerization occurs at two of any C2, C3 and C6 position, including combinations thereof, of the cyclodextrin moiety. For example, one cyclodextrin monomer precursor may be polymerized at two C6 positions while another cyclodextrin monomer precursor may be polymerized at a C2 and a C6 position of the cyclodextrin moiety. Using xcex2-cyclodextrin as an example, the lettering scheme for the relative position of each glucopyranose ring in a cyclodextrin is as follows: 
In a preferred embodiment of a linear cyclodextrin copolymer of the invention, the cyclodextrin monomer C has the following general formula (II): 
In formula (II), n and m represent integers which, along with the other two glucopyranose rings, define the total number of glucopyranose units in the cyclodextrin monomer. Formula (II) represents a cyclodextrin monomer which is capable of being polymerized at two C6 positions on the cyclodextrin unit. Examples of cyclodextrin monomers of formula (II) include, but are not limited to, 6A,6B-deoxy-xcex1-cyclodextrin (n=0, m=4), 6A,6C-deoxy-xcex1-cyclodextrin (n=1, m=3), 6A,6D-eoxy-xcex1-cyclodextrin (n=2, m=2), 6A,6B-deoxy-xcex2-cyclodextrin (n=0, m=5), 6A,6C-deoxy-xcex2cyclodextrin (n=1, m=4), 6A,6D-deoxy-xcex2-cyclodextrin (n=2, m=3), 6A,6B-deoxy-xcex3-cyclodextrin (n=0, m=6), 6A,6C-deoxy-xcex3-cyclodextrin (n=l, m=5), 6A,6D-deoxy-xcex3-cyclodextrin (n=2, m=4), and 6A,6E-deoxy-xcex3-cyclodextrin (n=3, m=3). In another preferred embodiment of linear cyclodextrin copolymer of the invention, a cyclodextrin monomer C unit has the following general formula (III): 
where p=5-7. In formula (III), one of D(+)-glucopyranose units of a cyclodextrin monomer has undergone ring opening to allow for polymerization at a C2 and a C3 position of the cyclodextrin unit. Cyclodextrin monomers of formula (III) are commercially available from Carbomer of Westborough, Mass. Examples of cyclodextrin monomers of formula (III) include, but are not limited to, 2A,3A-deoxy-2A,3A-dihydro-xcex1-cyclodextrin, 2A,3A-deoxy-2A,3A-dihydro-xcex2-cyclodextrin, 2A,3A-deoxy-2A,3A-dihydro-xcex3-cyclodextrin, commonly referred to as, respectively, 2,3-deoxy-xcex1-cyclodextrin, 2,3-deoxy-xcex2-cyclodextrin, and 2,3-deoxy-xcex3-cyclodextrin.
A comonomer A precursor may be any straight chain or branched, symmetric or asymmetric compound which upon reaction with a cyclodextrin monomer precursor, as described above, links two cyclodextrin monomers together. Preferably, a comonomer A precursor is a compound containing at least two functional groups through which reaction and thus linkage of the cyclodextrin monomers can be achieved. Examples of possible functional groups, which may be the same or different, terminal or internal, of each comonomer A precursor include, but are not limited to, amino, acid, ester, imidazole, and acyl halide groups and derivatives thereof. In a preferred embodiment, the two functional groups are the same and terminal. Upon copolymerization of a comonomer A precursor with a cyclodextrin monomer precursor, two cyclodextrin monomers may be linked together by joining the primary hydroxyl side of one cyclodextrin monomer with the primary hydroxyl side of another cyclodextrin monomer, by joining the secondary hydroxyl side of one cyclodextrin monomer with the secondary hydroxyl side of another cyclodextrin monomer, or by joining the primary hydroxyl side of one cyclodextrin monomer with the secondary hydroxyl side of another cyclodextrin monomer. Accordingly, combinations of such linkages may exist in the final copolymer. Both the comonomer A precursor and the comonomer A of the final copolymer may be neutral, cationic (e.g. by containing protonated groups such as, for example, quaternary ammonium groups) or anionic (e.g. by containing deprotonated groups, such as, for example, sulfate, phosphate or carboxylate anionic groups). The charge of comonomer A of the copolymer may be adjusted by adjusting pH conditions. Examples of suitable comonomer A precursors include, but are not limited to, cystamine, 1,6-diaminohexane, diimidazole, dithioimidazole, spermine, dithiospermine, dihistidine, dithiohistidine, succinimide (e.g. dithiobis(succinimidyl propionate) (DSP) and disuccinimidyl suberate (DSS)) and imidates (e.g. dimethyl 3,3xe2x80x2-dithiobispropionimidate (DTBP)). Copolymerization of a comonomer A precursor with a cyclodextrin monomer precursor leads to the formation of a linear cyclodextrin copolymer of the invention containing comonomer A linkages of the following general formulae:
xe2x80x94HNC(O)(CH2)xC(O)NHxe2x80x94, xe2x80x94HNC(O)(CH2)xSS(CH2)xC(O)NHxe2x80x94, xe2x80x94+H2N(CH2)xSS(CH2)xNH2+xe2x80x94, xe2x80x94HNC(O)(CH2CH2O)xCH2CH2C(O)NHxe2x80x94, xe2x95x90NNHC(O)(CH2CH2O)xCH2CH2C(O)NHNxe2x95x90, xe2x80x94+H2NCH2(CH2CH2O)xCH2CH2CH2NH2+xe2x80x94, xe2x80x94HNC(O)(CH2CH2O)xCH2CH2SS(CH2CH2O)xCH2CH2C(O)NHxe2x80x94, xe2x80x94HNC(NH2+)(CH2CH2O)xCH2CH2C(NH2+)NHxe2x80x94, xe2x80x94SCH2CH2NHC(NH2+)(CH2)xC(NH2+)NHCH2CH2Sxe2x80x94, xe2x80x94SCH2CH2NHC(NH2+)(CH2)xSS(CH2)xC(NH2+)NHCH2CH2Sxe2x80x94, xe2x80x94SCH2CH2NHC(NH2+)CH2CH2(OCH2CH2)xC(NH2+)NHCH2CH2Sxe2x80x94, 
In the above formulae, x=1-50, and y+z=x. Preferably, x=1-30. More preferably, x=1-20. In a preferred embodiment, comonomer A is biodegradable or acid-labile. Also in a preferred embodiment, the comonomer A precursor and hence the comonomer A may be selectively chosen in order to achieve a desired application. For example, to deliver small molecular therapeutic agents, a charged polymer may not be necessary and the comonomer A may be a polyethylene glycol group.
A linear cyclodextrin copolymer of the invention may be modified with at least one ligand attached to the cyclodextrin copolymer. The ligand may be attached to the cyclodextrin copolymer through the cyclodextrin monomer C or comonomer A. Preferably, the ligand is attached to at least one cyclodextrin moiety of the linear cyclodextrin copolymer. Preferably, the ligand allows a linear cyclodextrin copolymer to target and bind to a cell. If more than one ligand, which may be the same or different, is attached to a linear cyclodextrin copolymer of the invention, the additional ligand or ligands may be bound to the same or different cyclodextrin moiety or the same or different comonomer A of the copolymer. Examples of suitable ligands include, but are not limited to, vitamins (e.g. folic acid), proteins (e.g. transferrin, and monoclonal antibodies) and polysaccharides. The ligand will vary depending upon the type of delivery desired. For example, receptor-mediated delivery may by achieved by, but not limited to, the use of a folic acid ligand while antisense oligo delivery may be achieved by, but not limited to, use of a transferrin ligand. The ligand may be attached to a copolymer of the invention by means known in the art.
Another embodiment of the invention is a method of preparing a linear cyclodextrin copolymer. According to the invention, a linear cyclodextrin copolymer of the invention may be prepared by copolymerizing a cyclodextrin monomer precursor disubstituted with an appropriate leaving group with a comonomer A precursor capable of displacing the leaving groups. The leaving group, which may be the same or different, may be any leaving group known in the art which may be displaced upon copolymerization with a comonomer A precursor. In a preferred embodiment, a linear cyclodextrin copolymer may be prepared by iodinating a cyclodextrin monomer precursor to form a diiodinated cyclodextrin monomer precursor and copolymerizing the diiodinated cyclodextrin monomer precursor with a comonomer A precursor to form a linear cyclodextrin copolymer having a repeating unit of formula Ia, lb, or a combination thereof, each as described above. In a preferred embodiment, a method of preparing a linear cyclodextrin of the invention iodinates a cyclodextrin monomer precursor as described above to form a diiodinated cyclodextrin monomer precursor of formula IVa, IVb, IVc or a mixture thereof: 
The diiodinated cyclodextrin may be prepared by any means known in the art. (Tabushi et al. J. Am. Chem. 106, 5267-5270 (1984); Tabushi et al. J. Am. Chem. 106, 4580-4584 (1984)). For example, xcex2-cyclodextrin may be reacted with biphenyl-4,4xe2x80x2-disulfonyl chloride in the presence of anhydrous pyridine to form a biphenyl-4,4xe2x80x2-disulfonyl chloride capped xcex2-cyclodextrin which may then be reacted with potassium iodide to produce diiodo-xcex2-cyclodextrin. The cyclodextrin monomer precursor is iodinated at only two positions. By copolymerizing the diiodinated cyclodextrin monomer precursor with a comonomer A precursor, as described above, a linear cyclodextrin polymer having a repeating unit of formula Ia, Ib, or a combination thereof, also as described above, may be prepared. If appropriate, the iodine or iodo groups may be replaced with other known leaving groups.
Also according to the invention, the iodo groups or other appropriate leaving group may be displaced with a group that permits reaction with a comonomer A precursor, as described above. For example, a diiodinated cyclodextrin monomer precursor of formula IVa, IVb, IVc or a mixture thereof may be aminated to form a diaminated cyclodextrin monomer precursor of formula Va, Vb, Vc or a mixture thereof: 
The diaminated cyclodextrin monomer precursor may be prepared by any means known in the art. (Tabushi et al. Tetrahedron Lett. 18:1527-1530 (1977); Mungall et al., J. Org Chem. 1659-1662 (1975)). For example, a diiodo-xcex2-cyclodextrin may be reacted with sodium azide and then reduced to form a diamino-xcex2-cyclodextrin. The cyclodextrin monomer precursor is aminated at only two positions. The diaminated cyclodextrin monomer precursor may then be copolymerized with a comonomer A precursor, as described above, to produce a linear cyclodextrin copolymer having a repeating unit of formula Ia, Ib, or a combination thereof, also as described above. However, the amino functionality of a diaminated cyclodextrin monomer precursor need not be directly attached to the cyclodextrin moiety. Alternatively, the amino functionality may be introduced by displacement of the iodo or other appropriate leaving groups of a cyclodextrin monomer precursor with amino group containing moieties such as, for example, xe2x88x92SCH2CH2NH2, to form a diaminated cyclodextrin monomer precursor of formula Vd, Ve, Vf or a mixture thereof: 
A linear cyclodextrin copolymer of the invention may also be prepared by reducing a linear oxidized cyclodextrin copolymer of the invention as described below. This method may be performed as long as the comonomer A does not contain a reducible moiety or group such as, for example, a disulfide linkage.
According to the invention, a linear cyclodextrin copolymer of the invention may be oxidized so as to introduce at least one oxidized cyclodextrin monomer into the copolymer such that the oxidized cyclodextrin monomer is an integral part of the polymer backbone. A linear cyclodextrin copolymer which contains at least one oxidized cyclodextrin monomer is defined as a linear oxidized cyclodextrin copolymer. The cyclodextrin monomer may be oxidized on either the secondary or primary hydroxyl side of the cyclodextrin moiety. If more than one oxidized cyclodextrin monomer is present in a linear oxidized cyclodextrin copolymer of the invention, the same or different cyclodextrin monomers oxidized on either the primary hydroxyl side, the secondary hydroxyl side, or both may be present. For illustration purposes, a linear oxidized cyclodextrin copolymer with oxidized secondary hydroxyl groups has, for example, at least one unit of formula VIa or VIb: 
In formulae VIa and VIb, C is a substituted or unsubstituted oxidized cyclodextrin monomer and A is a comonomer bound, i.e. covalently bound, to the oxidized cyclodextrin C. Also in formulae VIa and VIb, oxidation of the secondary hydroxyl groups leads to ring opening of the cyclodextrin moiety and the formation of aldehyde groups.
A linear oxidized cyclodextrin copolymer may be prepared by oxidation of a linear cyclodextrin copolymer as discussed above. Oxidation of a linear cyclodextrin copolymer of the invention may be accomplished by oxidation techniques known in the art. (Hisamatsu et al., Starch 44:188-191 (1992)). Preferably, an oxidant such as, for example, sodium periodate is used. It would be understood by one of ordinary skill in the art that under standard oxidation conditions that the degree of oxidation may vary or be varied per copolymer. Thus in one embodiment of the invention, a linear oxidized copolymer of the invention may contain one oxidized cyclodextrin monomer. In another embodiment, substantially all to all cyclodextrin monomers of the copolymer would be oxidized.
Another method of preparing a linear oxidized cyclodextrin copolymer of the invention involves the oxidation of a diiodinated or diaminated cyclodextrin monomer precursor, as described above, to form an oxidized diiodinated or diaminated cyclodextrin monomer precursor and copolymerization of the oxidized diiodinated or diaminated cyclodextrin monomer precursor with a comonomer A precursor. In a preferred embodiment, an oxidized diiodinated cyclodextrin monomer precursor of formula VIIa, VIIb, VIIc, or a mixture thereof: 
may be prepared by oxidation of a diiodinated cyclodextrin monomer precursor of formulae IVa, IVb, IVc, or a mixture thereof, as described above. In another preferred embodiment, an oxidized diaminated cyclodextrin monomer precursor of formula VIIa, VIIIb, VIIIc or a mixture thereof: 
may be prepared by amination of an oxidized diiodinated cyclodextrin monomer precursor of formulae VIIa, VIIb, VIIc, or a mixture thereof, as described above. In still another preferred embodiment, an oxidized diaminated cyclodextrin monomer precursor of formula IXa, IXb, IXc or a mixture thereof: 
may be prepared by displacement of the iodo or other appropriate leaving groups of an oxidized cyclodextrin monomer precursor disubstituted with an iodo or other appropriate leaving group with the amino group containing moiety xe2x88x92SCH2CH2NH2.
Alternatively, an oxidized diiodinated or diaminated cyclodextrin monomer precursor, as described above, may be prepared by oxidizing a cyclodextrin monomer precursor to form an oxidized cyclodextrin monomer precursor and then diiodinating and/or diaminating the oxidized cyclodextrin monomer, as described above. As discussed above, the cyclodextrin moiety may be modified with other leaving groups other than iodo groups and other amino group containing functionalities. The oxidized diiodinated or diaminated cyclodextrin monomer precursor may then be copolymerized with a comonomer A precursor, as described above, to form a linear oxidized cyclodextrin copolymer of the invention.
A linear oxidized cyclodextrin copolymer may also be further modified by attachment of at least one ligand to the copolymer. The ligand is as described above.
According to the invention, a linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer may be attached to or grafted onto a substrate. The substrate may be any substrate as recognized by those of ordinary skill in the art. In another preferred embodiment of the invention, a linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer may be crosslinked to a polymer to form, respectively, a crosslinked cyclodextrin copolymer or a crosslinked oxidized cyclodextrin copolymer. The polymer may be any polymer capable of crosslinking with a linear or linear oxidized cyclodextrin copolymer of the invention (e.g. polyethylene glycol (PEG) polymer, polyethylene polymer). The polymer may also be the same or different linear cyclodextrin copolymer or linear oxidized cyclodextrin copolymer. Thus, for example, a linear cyclodextrin copolymer may be crosslinked to any polymer including, but not limited to, itself, another linear cyclodextrin copolymer, and a linear oxidized cyclodextrin copolymer. A crosslinked linear cyclodextrin copolymer of the invention may be prepared by reacting a linear cyclodextrin copolymer with a polymer in the presence of a crosslinking agent. A crosslinked linear oxidized cyclodextrin copolymer of the invention may be prepared by reacting a linear oxidized cyclodextrin copolymer with a polymer in the presence of an appropriate crosslinking agent. The crosslinking agent may be any crosslinking agent known in the art. Examples of crosslinking agents include dihydrazides and disulfides. In a preferred embodiment, the crosslinking agent is a labile group such that a crosslinked copolymer may be uncrosslinked if desired.
A linear cyclodextrin copolymer and a linear oxidized cyclodextrin copolymer of the invention may be characterized by any means known in the art. Such characterization methods or techniques include, but are not limited to, gel permeation chromatography (GPC), matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF Mass spec), 1H and 13C NMR, light scattering and titration.
The invention also provides a cyclodextrin composition containing at least one linear cyclodextrin copolymer and at least one linear oxidized cyclodextrin copolymer of the invention as described above. Accordingly, either or both of the linear cyclodextrin copolymer and linear oxidized cyclodextrin copolymer may be crosslinked to another polymer and/or bound to a ligand as described above. Therapeutic compositions according to the invention contain a therapeutic agent and a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer, including crosslinked copolymers, of the invention. A linear cyclodextrin copolymer, a linear oxidized cyclodextrin copolymer and their crosslinked derivatives are as described above. The therapeutic agent may be any synthetic or naturally occurring biologically active therapeutic agent including those known in the art. Examples of suitable therapeutic agents include, but are not limited to, antibiotics, steroids, polynucleotides (e.g. genomic DNA, cDNA, mRNA and antisense oligonucleotides), plasmids, peptides, peptide fragments, small molecules (e.g. doxorubicin) and other biologically active macromolecules such as, for example, proteins and enzymes.
A therapeutic composition of the invention may be prepared by means known in the art. In a preferred embodiment, a copolymer of the invention is mixed with a therapeutic agent, as described above, and allowed to self-assemble. According to the invention, the therapeutic agent and a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer of the invention associate with one another such that the copolymer acts as a delivery vehicle for the therapeutic agent. The therapeutic agent and cyclodextrin copolymer may associate by means recognized by those of skill in the art such as, for example, electrostatic interaction and hydrophobic interaction. The degree of association may be determined by techniques known in the art including, for example, fluorescence studies, DNA mobility studies, light scattering, electron microscopy, and will vary depending upon the therapeutic agent. As a mode of delivery, for example, a therapeutic composition of the invention containing a copolymer of the invention and DNA may be used to aid in transfection, i.e. the uptake of DNA into an animal (e.g. human) cell. (Boussif, O. Proceedings of the National Academy of Sciences, 92:7297-7301 (1995); Zanta et al. Bioconjugate Chemistry, 8:839-844 (1997)).
A therapeutic composition of the invention may be, for example, a solid, liquid, suspension, or emulsion. Preferably a therapeutic composition of the invention is in a form that can be injected intravenously. Other modes of administration of a therapeutic composition of the invention include, depending on the state of the therapeutic composition, methods known in the art such as, but not limited to, oral administration, topical application, parenteral, intravenous, intranasal, intraocular, intracranial or intraperitoneal injection.
Depending upon the type of therapeutic agent used, a therapeutic composition of the invention may be used in a variety of therapeutic methods (e.g. DNA vaccines, antibiotics, antiviral agents) for the treatment of inherited or acquired disorders such as, for example, cystic fibrosis, Gaucher""s disease, muscular dystrophy, AIDS, cancers (e.g., multiple myeloma, leukemia, melanoma, and ovarian carcinoma), cardiovascular conditions (e.g., progressive heart failure, restenosis, and hemophilia), and neurological conditions (e.g., brain trauma). According to the invention, a method of treatment administers a therapeutically effective amount of a therapeutic composition of the invention. A therapeutically effective amount, as recognized by those of skill in the art, will be determined on a case by case basis. Factors to be considered include, but are not limited to, the disorder to be treated and the physical characteristics of the one suffering from the disorder.
Another embodiment of the invention is a composition containing at least one biologically active compound having agricultural utility and a linear cyclodextrin copolymer or a linear oxidized cyclodextrin copolymer of the invention. The agriculturally biologically active compounds include those known in the art. For example, suitable agriculturally biologically active compounds include, but are not limited to, fungicides, herbicides, insecticides, and mildewcides.